Directed evolution of oxidase enzymes

C - Chemistry – Metallurgy – 12 – N

Patent

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Details

C12N 9/00 (2006.01) C07H 21/04 (2006.01) C12N 9/04 (2006.01) C12N 15/00 (2006.01) C12N 15/10 (2006.01)

Patent

CA 2408060

This invention relates to the expression of improved polynucleotide and polypeptide sequences encoding for eukaryotic enzymes, particularly oxidase enzymes. The enzymes are advantageously produced in conventional or facile expression systems. Various methods for directed evolution of polynucleotide sequences can be used to obtain the improved sequences. The improved characteristics of the polypeptides or proteins generated in this manner include improved expression, enhanced activity toward one or more substrates, and increased thermal stability. In a particular embodiment, the invention relates to improved expression of the galactose oxidase gene and galactose oxidase enzymes. GAO mutants that are highly active and/or thermostable are disclosed.

L'invention concerne l'expression de séquences polynucléotidiques et polypeptidiques améliorées codant pour les enzymes eucariotes, plus particulièrement les enzymes oxydases. Ces enzymes sont produites de manière avantageuse dans des systèmes d'expression faciles ou classiques. Il est possible d'utiliser divers procédés d'évolution dirigée de séquences polynucléotidiques pour obtenir les séquences améliorées. Les polypeptides ou les protéines élaborés selon ce procédé comprennent, entre autres caractéristiques améliorées, une expression améliorée, une activité améliorée sur un ou plusieurs substrats, et une plus grande stabilité thermique. Un mode de réalisation décrit dans l'invention concerne l'expression améliorée du gène de la galactose oxydase et des enzymes de la galactose oxydase. L'invention concerne également des mutants de la galactose oxydase présentant une activité élevée et/ou une thermostabilité élevée.

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